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TDP-43-Mediated Toxicity in HEK293T Cells: A Fast and Reproducible Protocol To Be Employed in the Search of New Therapeutic Options against Amyotrophic Lateral Sclerosis
Cytoplasmic TDP-43 aggregates are a hallmark of amyotrophic lateral sclerosis (ALS). Today, only two drugs are available for ALS treatment, and their modest effect prompts researchers to search for new therapeutic options. TDP-43 represents one of the most promising targets for therapeutic intervent...
Autores principales: | , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2019
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7016571/ https://www.ncbi.nlm.nih.gov/pubmed/31888078 http://dx.doi.org/10.3390/cells9010068 |
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author | Lanznaster, Débora Bourgeais, Jérôme Bruno, Clement Hergesheimer, Rudolf C. Thepault, Rose-Anne Vourc’h, Patrick Corcia, Philippe Andres, Christian R. Herault, Olivier Blasco, Hélène |
author_facet | Lanznaster, Débora Bourgeais, Jérôme Bruno, Clement Hergesheimer, Rudolf C. Thepault, Rose-Anne Vourc’h, Patrick Corcia, Philippe Andres, Christian R. Herault, Olivier Blasco, Hélène |
author_sort | Lanznaster, Débora |
collection | PubMed |
description | Cytoplasmic TDP-43 aggregates are a hallmark of amyotrophic lateral sclerosis (ALS). Today, only two drugs are available for ALS treatment, and their modest effect prompts researchers to search for new therapeutic options. TDP-43 represents one of the most promising targets for therapeutic intervention, but reliable and reproducible in vitro protocols for TDP-43-mediated toxicity are lacking. Here, we used HEK293T cells transfected with increasing concentrations of TDP-43-expressing plasmid to evaluate different parameters of toxicity and alterations in cellular metabolism. Overexpression of TDP-43 induced aggregates occurrence followed by the detection of 25- and 35-kDa forms of TDP-43. TDP-43 overexpression decreased cell viability and increased cells arrested at G2/M phase and nuclear fragmentation. Analysis of the energetic metabolism showed a tendency to decrease oxidative phosphorylation and increase glycolysis, but no statistical differences were observed. Metabolomics revealed alterations in different metabolites (mainly sphingolipids and glycerophospholipids) in cells overexpressing TDP-43. Our data reveal the main role of TDP-43 aggregation in cellular death and highlight novel insight into the mechanism of cellular toxicity induced by TDP-43. Here, we provide a simple, sensitive, and reliable protocol in a human-derived cell line to be used in high-throughput screenings of potential therapeutic molecules for ALS treatment. |
format | Online Article Text |
id | pubmed-7016571 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2019 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-70165712020-03-04 TDP-43-Mediated Toxicity in HEK293T Cells: A Fast and Reproducible Protocol To Be Employed in the Search of New Therapeutic Options against Amyotrophic Lateral Sclerosis Lanznaster, Débora Bourgeais, Jérôme Bruno, Clement Hergesheimer, Rudolf C. Thepault, Rose-Anne Vourc’h, Patrick Corcia, Philippe Andres, Christian R. Herault, Olivier Blasco, Hélène Cells Article Cytoplasmic TDP-43 aggregates are a hallmark of amyotrophic lateral sclerosis (ALS). Today, only two drugs are available for ALS treatment, and their modest effect prompts researchers to search for new therapeutic options. TDP-43 represents one of the most promising targets for therapeutic intervention, but reliable and reproducible in vitro protocols for TDP-43-mediated toxicity are lacking. Here, we used HEK293T cells transfected with increasing concentrations of TDP-43-expressing plasmid to evaluate different parameters of toxicity and alterations in cellular metabolism. Overexpression of TDP-43 induced aggregates occurrence followed by the detection of 25- and 35-kDa forms of TDP-43. TDP-43 overexpression decreased cell viability and increased cells arrested at G2/M phase and nuclear fragmentation. Analysis of the energetic metabolism showed a tendency to decrease oxidative phosphorylation and increase glycolysis, but no statistical differences were observed. Metabolomics revealed alterations in different metabolites (mainly sphingolipids and glycerophospholipids) in cells overexpressing TDP-43. Our data reveal the main role of TDP-43 aggregation in cellular death and highlight novel insight into the mechanism of cellular toxicity induced by TDP-43. Here, we provide a simple, sensitive, and reliable protocol in a human-derived cell line to be used in high-throughput screenings of potential therapeutic molecules for ALS treatment. MDPI 2019-12-26 /pmc/articles/PMC7016571/ /pubmed/31888078 http://dx.doi.org/10.3390/cells9010068 Text en © 2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Lanznaster, Débora Bourgeais, Jérôme Bruno, Clement Hergesheimer, Rudolf C. Thepault, Rose-Anne Vourc’h, Patrick Corcia, Philippe Andres, Christian R. Herault, Olivier Blasco, Hélène TDP-43-Mediated Toxicity in HEK293T Cells: A Fast and Reproducible Protocol To Be Employed in the Search of New Therapeutic Options against Amyotrophic Lateral Sclerosis |
title | TDP-43-Mediated Toxicity in HEK293T Cells: A Fast and Reproducible Protocol To Be Employed in the Search of New Therapeutic Options against Amyotrophic Lateral Sclerosis |
title_full | TDP-43-Mediated Toxicity in HEK293T Cells: A Fast and Reproducible Protocol To Be Employed in the Search of New Therapeutic Options against Amyotrophic Lateral Sclerosis |
title_fullStr | TDP-43-Mediated Toxicity in HEK293T Cells: A Fast and Reproducible Protocol To Be Employed in the Search of New Therapeutic Options against Amyotrophic Lateral Sclerosis |
title_full_unstemmed | TDP-43-Mediated Toxicity in HEK293T Cells: A Fast and Reproducible Protocol To Be Employed in the Search of New Therapeutic Options against Amyotrophic Lateral Sclerosis |
title_short | TDP-43-Mediated Toxicity in HEK293T Cells: A Fast and Reproducible Protocol To Be Employed in the Search of New Therapeutic Options against Amyotrophic Lateral Sclerosis |
title_sort | tdp-43-mediated toxicity in hek293t cells: a fast and reproducible protocol to be employed in the search of new therapeutic options against amyotrophic lateral sclerosis |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7016571/ https://www.ncbi.nlm.nih.gov/pubmed/31888078 http://dx.doi.org/10.3390/cells9010068 |
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